In renal proximal tubule (PT) cells, the pivotal event in serum phosphate regulation is the insertion, retention and recovery of the sodium-phosphate 2a (NaPiIIa) co-transporter at the apical membrane. NaPiIIa activity is influence by its direct interaction with a number of distinct PDZ proteins, including EBP50 and PDZK1. These proteins appear to influence the delivery and retention of NaPiIIa in the apical membrane. More recently, a novel PDZ protein, Shank2E, was implicated in regulating NaPiIIa. Already known to bind endocytic proteins (e.g. dynamin, cortactin), Shank2E binds NaPiIIa and redistribute with NaPiIIa into the cell interior when NaPiIIa was endocytosed from the apical membrane. Consequently, the following proposal will test the hypothesis that Shank2E coordinates the endocytosis of NaPiIIa in a regulated manner. To this end, the proposed studies will (1) define the regulated protein-protein interactions between Shank2E and NaPiIIaassociated proteins, (2) determine the role of Shank2E in moderating NaPiIIa activity in proximal tubule cell models and (3) demonstrate the role of Shank2E in NaPiIIa regulation in whole animal models. Results from these studies will make important advances on three fronts. At the molecular level, the studies will determine how modifications in Shank2E-centered protein-protein interactions with other PDZ domain proteins and NaPiIIa are coordinately regulated. At the cellular and organ level, the studies will elucidate how these coordinately regulated interactions result in Shank2E directing the endocytic recovery of NaPiIIa in response to physiologic stimuli. Finally, these results may provide mechanistic insight into phosphatemic diseases and direct the genesis of novel therapeutic strategies. PUBLIC HEALTH RELEVANCE: The broad goal of the proposed studies is to better understand how the body regulates the movement of salts and water in and out of the body. The kidney is the central organ involved in maintaining salt and water balance in the body. The balance of phosphate by the kidney is essential for maintaining the health status of the body. Chronic decreases of phosphate levels can result in weaknesses and deformities in bones and teeth. Alternatively, even modest increases in phosphate levels can result in vascular deposits and cardiovascular disease. The pivotal event in regulating the balance of phosphate in the body is the activity level of a phosphate transport protein in the kidney. The proposed studies seek to discover and define how a recently identified protein in the kidney (termed Shank2E) is capable of turning down the activity of the phosphate transport protein. Interestingly, the Shank2E protein also binds other key transport proteins. These include a chloride channel that is linked to Cystic Fibrosis and a bilirubin transport protein in the liver that is linked to Dubin-Johnson Syndrome. By discovering how Shank2E functions to regulate these proteins the findings elucidate how the body maintains itself under normal conditions, how changes in these proteins might give rise to specific diseases and how researchers might take advantage of these regulated mechanisms to develop medical therapies to treat these diseases.